Original Research ARTICLE
Enhancer associated long non-coding RNA transcription and gene regulation in experimental models of rickettsial infection
- 1The University of Texas Medical Branch at Galveston, United States
Recent discovery that much of the mammalian genome does not encode protein-coding genes (PCGs) has brought widespread attention to long noncoding RNAs (lncRNAs) as a novel layer of biological regulation. Enhancer lnc (elnc) RNAs from the enhancer regions of the genome carry the capacity to regulate PCGs in cis or in trans. Spotted fever rickettsioses represent the consequence of host infection with Gram-negative, obligate intracellular bacteria in the Genus Rickettsia. Despite being implicated in the pathways of infection and inflammation, the roles of lncRNAs in host response to Rickettsia species have remained a mystery. We have profiled the expression of host lncRNAs during infection of susceptible mice with R. conorii as a model closely mimicking the pathogenesis of human spotted fever rickettsioses. RNA sequencing on the lungs of infected hosts yielded reads mapping to 74,964 non-coding RNAs, 206 and 277 of which were determined to be significantly up- and down-regulated, respectively, in comparison to uninfected controls. Following removal of short non-coding RNAs and ambiguous transcripts, remaining transcripts underwent in-depth analysis of mouse lung epigenetic signatures H3K4Me1 and H3K4Me3, active transcript markers (POLR2A, p300, CTCF), and DNaseI hypersensitivity sites to identify two potentially active and highly up-regulated elncRNAs NONMMUT013718 and NONMMUT024103. Using Hi-3C sequencing resource, we further determined that genomic loci of NONMMUT013718 and NONMMUT024103 might interact with and regulate the expression of nearby PCGs, namely Id2 (inhibitor of DNA binding 2) and Apol10b (apolipoprotein 10b), respectively. Heterologous reporter assays confirmed the activity of elncRNAs as the inducers of their predicted PCGs. In the lungs of infected mice, expression of both elncRNAs and their targets was significantly higher than mock-infected controls. Induced expression of NONMMUT013718/Id2 in murine macrophages and NONMMUT024103/Apol10b in endothelial cells was also clearly evident during R. conorii infection in vitro. Finally, shRNA mediated knock-down of NONMMUT013718 and NONMMUT024103 elncRNAs resulted in reduced expression of endogenous Id2 and Apl10b, demonstrating the regulatory roles of these elncRNAs on their target PCGs. Our results provide very first experimental evidence suggesting altered expression of pulmonary lncRNAs and elncRNA-mediated regulation of PCGs involved in immunity and during host interactions with pathogenic rickettsiae.
Keywords: Long non-coding (lnc) RNA, Rickettsia, Enhancer long non-coding (elnc) RNA, RNA sequencing, transcription start site, Inhibitor of DNA binding 2 protein, Apolipoprotein 10b, host immune responses
Received: 24 Jul 2018;
Accepted: 05 Dec 2018.
Edited by:Adam Williams, Jackson Laboratory, United States
Reviewed by:Michael F. Minnick, The University of Montana, United States
Kushagra Bansal, Harvard Medical School, United States
Copyright: © 2018 Sahni, Chowdhury, Narra, Sahni, Khanipov and Fofanov. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
Dr. Sanjeev K. Sahni, The University of Texas Medical Branch at Galveston, Galveston, United States, firstname.lastname@example.org
Dr. Imran H. Chowdhury, The University of Texas Medical Branch at Galveston, Galveston, United States, email@example.com